This invention relates to a method for stimulating the cells of the immune system of a human by administering human growth hormone. More particularly, the invention relates to a method for increasing the total lymphocyte count of a human by administering human growth hormone. Even more particularly, the invention relates to a method for increasing the T4/T8 ratio in a human by administering human growth hormone.
2. Background Art
The immune systems of vertebrates generally provide a dual mechanism for recognizing and eliminating materials which are foreign to the body. These mechanisms, which are ordinarily referred to as humoral (antibody mediated) and cellular immunity, are provided by B- and T- lymphocytes, respectively.
The immune response to foreign material, or antigens, involves complex interactions of various lymphocytes, or B-and T-cells. These interactions can be severely compromised following certain infections and after injury. T-cells, which are derived from the thymus, are particularly important for the maintenance of immunocompetence following injury (Hoyt, et al., J. of Trauma, Vol. 30, p. 759, 1990).
There are several subsets of T-cells which are named by their functions. Helper T-cells (T.sub.H) are involved in the activation of B-cells into plasma cells which produce antibodies, which in turn, react with foreign antigens. T.sub.H cells express a surface antigen called CD4 or T4. T4.sup.+ cells comprise predominantly T.sub.H cells, along with another type of T-cell, called the accessory T-cell (T.sub.A). Cytotoxic T-cells (T.sub.C) are involved in the destruction of antigen-bearing cells. Another cell, derived from the same precursor as the T-cell, the natural killer (NK) cell, aids the T.sub.C in protecting the body from tumor cells. T.sub.C cells express a different surface antigen from T.sub.H cells which is called CD8 or T8. T8.sup.+ cells consist of T-cells with both suppressor (T.sub.S) and cytotoxic activities. T-cell function and a normal T4/ T8 ratio are important to providing cellular immunity, which, in particular, defends the body from foreign antigens including invaders such as bacteria, fungi, parasites, and viruses.
The ratio of T4.sup.+ to T8.sup.+ cells is tightly regulated in humans, and has recently been shown to decrease as a result of severe head injury (See e.g. Hoyt, et al, 1990). This decreased T4/T8 ratio may result from alterations in the production of various cytokines, alterations to macrophages and their metabolites, and by activation of T.sub.S cells which inhibit the proliferation of other T-cells. Activation of T.sub.S cells probably may occur as a result of injury-induced serum suppressor substances such as prostaglandin E.sub.2 (PGE.sub.2), leukotrienes, tumor necrosis factor, and lymphocyte and proteolytic fragments of cytokines and their receptors.
A decreased T4/T8 ratio has also been shown to be characteristic of patients having acquired immune deficiency syndrome (AIDS) which is caused by infection with human immunodeficiency virus (HIV; Cease, et al., in AIDS Vaccine Research and Clinical Trials, Putney and Bolognesi, eds., p. 139, 1990). In the case of AIDS, one mechanism for the decreased T4/T8 ratio is the utilization of the T4 molecule by HIV as a cellular receptor. In binding to the T4 molecule, the virus is provided entry to that particular cell, which ultimately results in disablement of the normal cellular functions. The decrease in circulating T4.sup.+ lymphocytes is caused not only by the death of these cells following infection, but also from the loss of cell surface expression of T4 molecules on HIV infected T4.sup.+ cells (Orloff and McDougal, in AIDS Vaccine Research and Clinical Trials, Putney and Bolognesi, eds., p. 63, 1990). Infection by other viruses, which do not use the T4 molecule as a receptor, may result in immune suppression by other mechanisms which are not all completely understood. Oncogenic or tumor forming viruses can cause immunosuppression by inducing cells to form tumors which release immunosuppressive factors. Other viruses, such as cytomegalovirus (CMV) and lactic dehydrogenase virus (LDV) inhibit cellular and humoral immunity by unknown mechanisms.
HGH is a 191 amino acid single chain protein which is released by the anterior pituitary. It has a molecular weight of 21,500 kilodaltons and has disulfide bonds linking amino acids 53 and 165 and amino acids 182 and 189 (Niall, Nature New Biol. Vol. 230, p. 90, 1977). HGH is a potent anabolic agent, especially due to retention of nitrogen, phosphorus, potassium and calcium.
HGH causes a variety of physiological and metabolic effects in various animal models including linear bone growth, lactation, activation of macrophages, insulin-like and diabetogenic effects (Chawla, et al., Ann. Rev. Med. Vol. 34, p. 519, 1983; Isaksson, et al., Ann. Rev. Physiol., Vol. 47, p. 483, 1985; Edwards, et al. Science Vol. 239, p. 769, 1988; Thorner and Vance, J. Clin. Invest. Vol. 82, p. 745, 1988; Hughes and Friesen, Ann. Rev. Physiol. Vol. 47, p. 469, 1985). Treatment of hypophysectomized rats with GH can restore at least a portion of the growth rate of the rats (Moore, et al., Endocrinology, Vol. 122, p. 2920, 1988).
A connection has been said to exist between the anterior pituitary and the immune system, and specifically with GH. Human growth hormone (HGH) is believed to be necessary for maintaining lymphoid tissues populated with lymphocytes. In mice, GH removal results in thymic atrophy, which can be reversed by readministration of GH (Baroni, Experientia, Vol. 23, p. 282, 1967). Two groups of investigators concluded from their studies that GH controls the growth of lymphoid tissue (Pierpaoli and Sorkin, Nature, Vol. 215, p. 834, 1967; Baroni, Experientia, Vol. 23, (1967). Subsequently, immunologic function was restored in the pituitary dwarf mouse by a combination of bovine somatotropic hormone and thyroxin (Baroni et al., Immunol., Vol. 17, p. 303, 1969).
In a sex-linked dwarf chicken strain, bovine GH treatment resulted in enhanced antibody responses and bursal growth while thyroxine treatment stimulated thymus growth (Marsh et al., Proc. Soc. Exp. Biol. Med., Vol. 175, p. 351, 1984). However, neither treatment altered immune function in the autosomal dwarf chicken. Bovine GH therapy alone partially restored immunologic function in immunodeficient Weimaraner dogs (Roth et al., Ann. J. Vet. Res., Vol. 45, p. 1151, 1984).
Mice with hereditary GH deficiency develop an impairment of the immune system associated with thymic atrophy, immunodeficiency, and wasting, resulting in a shortened life expectancy (Frabris et al., Clin. Exp. Immunol., Vol 9, p. 209, 1971). It has been shown that an age-associated decline in the plasma concentration of thymulin (a thymic hormone) occurs and that plasma thymulin concentration increases in bGH-treated middle-aged and old dogs (Goff et al., Clin. Exp. Immunol., Vol 68, p. 580, 1987). Further, administration of HGH to C.sub.57 /B1/6J mice was found to reverse the inhibitory effect of prednisolone on thymus and spleen cellularity and on natural killer activity; administration of HGH without prednisolone had no effect, although at higher doses it induced a decrease of thymic parameters and natural killer activity with no effect on spleen cellularity, and relative weights (Franco et al., Acta Endocrinologica, Vol. 123, p. 339, 1990).
Two different human receptors have been cloned with which HGH appears to interact; the HGH liver receptor (Leung et al., Nature, Vol. 330, p. 537, 1987) and the human prolactin receptor (Boutin et al., Mol. Endocrinol. Vol. 3, p. 1455, (1989). However, there may be others including the human placental lactogen receptor (Freemark, et al., Endocrinol. Vol. 120, p. 1865, 1987). These homologous receptors contain a glycosylated extracellular hormone binding domain, a single transmembrane domain and a cytoplasmic domain which differs considerably in sequence and size. One or more receptors play a role in determining the physiological response to HGH.
It has been reported that, especially in women after menopause, GH secretion declines with age. Millard et al., Neurobiol, Aging, Vol. 11, p. 229, 1990; Takahashi et al., Neuroendocrinology, Vol. 46, p. 137, 1987). See also Rudman et al., J. Clin. Invest., Vol. 67, p. 1361, 1981 and Blackman, Endocrinology and Aging, Vol. 16, p. 981, 1987. Moreover, a report exists that some of the manifestations of aging, including decreased lean body mass, expansion of adipose-tissue mass, and the thinning of the skin, may be reduced by GH treatment three times a week. See, e.g., Rudman et al., N. Eng. J. Med., Vol. 323, p. 1, 1990 and the accompanying article in the same journal issue by Dr. Vance (pp. 52-54).
HGH is released in response to stimulation by human growth hormone releasing hormone, hGHRH, which is released by the hypothalamus. hGHRH is also referred to as human growth hormone releasing factor, hGHRF or GRF, as somatoliberin or as somatocrinin. HGH stimulates the growth of many tissues of the body, exerting many of its effects by stimulating the secretion of other growth factors, such as the somatomedins, which display insulin-like activities (U.S. Pat. No. 4,769,361). A major biological effect of HGH is to promote growth in young mammals and to maintain tissues in older mammals. The organ systems affected include the skeleton, connective tissue, muscles, and viscera such as liver, intestine, and kidneys. Growth hormone exerts its effect through interaction with specific receptors on the target cell's membrane. HGH is a member of a family of homologous hormones that include placental lactogens, prolactins, and other genetic and species variants of growth hormone (Nicoll, et al., Endocrine Reviews, Vol. 7, p. 169, 1986). HGH is unusual among these in that it exhibits broad species specificity and binds to either the cloned somatogenic (Leung, et al. Nature, Vol. 330, p. 537, 1987) or prolactin receptor (Boutin, et al., Cell, Vol. 53, p. 69, 1988). The cloned gene for HGH has been expressed in secreted form in E. coli (Chang, et al. (11987) Gene, Vol. 39, p. 247, 1987). The effects of HGH include linear growth (somatogenesis), lactation, activation of macrophages, and other insulin-like and diabetogenic effects (Chawla, Ann. Rev. Med., Vol. 34, p. 519, 1983; Edwards, et al., Science, Vol. 239, p. 769, 1988; Thorner, et al., J. Clin. Invest., Vol. 81, p. 745, 1988).
HGH has been used primarily in the treatment of hypopituitary dwarfism (Rapaport, et al., J. of Pediatrics, Volume 109, p. 434, 1986). HGH treatment in growth hormone deficient patients results in the stimulation of skeletal growth, an increase in cellular protein synthesis, an increase in serum glucose and insulin levels, a reduction in body fat stores, and stimulation of connective tissue and mineral metabolism. Among its most striking effects in hypopituitary (GH-deficient) subjects is accelerated linear bone growth of bone-growth-plate-cartilage resulting in increased stature (Kaplan, Growth Disorders in Children and Adolescents, Springfield, Ill., Charles C. Thomas, 1964).
In addition to being used to stimulate growth, HGH has also been used as a dietary supplement to maintain a positive nitrogen balance (U.S. Pat. No. 4,863,901), for the treatment of intoxicated individuals (U.S. Pat. No. 4,816,439) and for the treatment and diagnosis of neurodegenerative diseases such as Alzheimer's Disease and Parkinson's Disease (U.S. Pat. Nos. 4,939,124 and 4,791,099). Growth hormone (obtained from either rats or pigs) has also been shown to act on the immune system of animals by increasing the production of macrophages, and by activating their oxidative metabolism (U.S. Pat. No. 4,837,202).
Studies have also recently been conducted on GH and T-cell proliferation in the thymus (Murphy et al., FASEB Meeting Abstract, Atlanta, April 1991; Durum et al., FASEB Meeting Abstract, Atlanta, April 1991). For other articles on the immune effects of GH, see Kelley, "Growth Hormone in Immunobiology," in Psychoneuroimmunology II, 2nd Ed., B. Ader et al., eds., Acad. Press 1990, and Ammann, "Growth Hormone and Immunity," in Human Growth Hormone--Progress and Challenges, L. Underwood, ed., Marcel Dekker, Inc., New York, p. 243, 1988; and Weigent and Blalock, Prog. NeuroEndocrinImmunology, Vol 3, p. 231, 1990; and Kelly, Biochem. Pharmacol., Vol. 38, p. 705, 1989.
Human growth hormone treatment has been shown to suppress some immunological functions in growth hormone-deficient children (Rapaport, et al., 1986, Bozzola, et al., Acta. Paediatr. Scand., Vol. 74, p. 675, 1988). However, there is a difference in the responses of lymphocytes from growth hormone-deficient patients and normal controls to HGH. Rapaport et al. reported that when HGH was administered to normal subjects, a significant depression in spontaneous lymphocyte proliferation was seen. In contrast, a significant increase in proliferation was demonstrated when HGH was administered to the growth hormone deficient group. (Rapaport, et al., Life Sciences, Vol. 41, p. 231, 1987). It therefore appears that HGH has different effects on immune function, depending on the status of the patient's hormonal interactions and baseline immune responsiveness. Some studies suggest that administration of HGH to HGH deficient patients decreases the T4/T8 ratio (Rapaport, et al., 1986).